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  1. May 2020
    1. For morphometric comparison of human islets from 12-LO-positive and -negative cases, a composite image of 50 visual fields at ×200 magnification (area of 50 000 μm2) was captured using the mosaic function of AxioVision analysis software (v4.7; Zeiss). Five composite images of the pancreas were obtained per case, and at least 3 cases per group were studied. Areas positive for insulin or 12-LO were defined by setting a threshold signal for each antigen, and the size of the positive area was quantified automatically based on this threshold. Hematoxylin and eosin (HE)-stained images on the nPOD website from serial sections of the blocks we analyzed were used to determine the islet area for each donor. To quantitate the PP staining intensity, images of 30 islets from the uncinate and nonuncinate areas of pancreas sections immunostained for PP were analyzed for mean densitometric values of PP+ areas using AxioVision v4.7. To normalize staining, mean densitometric background staining with nonimmune serum was subtracted from values obtain for the PP sections. Observers blinded to the clinical information of donors performed all measurements

      12-LO staining and analysis